As a result of the sequencing of the human genome, it has become apparent that the complexity of an organism does not necessarily correlate with the size of its genome. This has sparked an interest in discovering exactly how genes are regulated on levels other than primary genomic sequence. Epigenetics has arisen as a field that addresses this concern by focusing on post-transcriptional methods of gene expression control including DNA methylation, histone post-translational modifications (PTMs) and non-coding RNAs (1). Modifications made to proteins post-translationally can affect their function, location or longevity. One set of highly modified proteins important to gene expression are histones. DNA is wrapped around histones, which form nucleosomes before being condensed into chromatin and ultimately chromosomes.
Histone PTMs have been shown to be very important to gene expression. Some modifications serve to signal the recruitment of chromatin modifying enzymes while some serve to alter the interaction between the histones and DNA allowing or prohibiting the access of transcription machinery (2). Some more common histone modifications include methylation, acetylation and phosphorylation. Of these common modifications, methylation is, by far, the most complex. Both lysine and arginine residues can be modified by mono- or di-methylation and lysine residues can be tri-methylated as well. These varying states of methylation can be associated with both active and inactive genes. The complexity and importance of methylation, on histones in particular, has stirred much interest in the enzymes capable of adding (methyltransferases) and removing (demethylases) these modifications.
The first demethylase was discovered in 2004 and was termed Lysine Specific Demethylase 1 or LSD1 (3). LSD1 is a flavin-dependent amine oxidase that can remove mono- and di-methyl marks from H3K4 primarily, H3K9 under certain conditions and some non-histone substrates such as p53 (4-6). It has been shown to be part of many protein complexes including CoREST, NuRD, and AR/ER (7). LSD1 is also associated with gene repression and has been suggested to be important in initiating myc-induced transcription in cancers (3, 8-11).
Structural and biochemical studies have led to the development of numerous LSD1 inactivators that have the potential to be therapeutic tools, much like the successful deacetylase inhibitors currently in use (12). In addition, the mechanism of LSD1 indicates that it is an excellent candidate for suicide inactivators. Many monoamine oxidase (MAO) inhibitors have been suggested as potential LSD1 suicide inactivators (13). Several different assays are used to study the activity of LSD1 in the presence and absence of these various inhibitors in order to determine their efficiency.
With the importance of histone methylation and demethylation in normal gene regulation and aberrant gene regulation in cancer and other diseases, improved tools to characterize the activity and specificity of enzymes catalyzing methylation and demethylation of histones and other proteins are needed.